Accumulation of 13C-labeled phosphatidylethanolamine in the termite abdomen revealed by correlative isotope microscopy and mass microscopy.

Isotope microscopic imaging and atmospheric pressure matrix-assisted laser desorption/ionization mass spectrometry imaging (AP-MALDI-MSI) provide powerful, complementary approaches for visualizing metabolic dynamics in biological tissues. This study applied these techniques to termite workers fed with 13C-labeled cellulose for 1 week. Termites are classified as eusocial insects because of their colonies' clear division of labor. The two primary castes in their life cycle are reproductive (king and queen), responsible for reproduction, and non-reproductive (workers and soldiers), who handle tasks such as defense, brood care and foraging. Although various techniques have been developed to detect 13C-labeled biomolecules in samples, it remains unclear whether the iMScopeTM prototype can visualize these molecules with high spatial resolution. Advanced isotope microscopic imaging technique with high spatial resolution (200-300 nm) offered ultra-high-resolution visualization of the relative abundance of the 13C/12C distribution, suggesting precise localization of isotope enrichment in the abdomen. AP-MALDI-MSI performed in the iMScopeTM prototype enabled spatial mapping of 13C-labeled and unlabeled metabolites, such as acetyl-L-carnitine and phosphatidylethanolamine (PE), by detecting characteristic mass shifts due to 13C incorporation. The accumulation of PE in the termite abdomen represents an adaptive strategy to optimize nutrient allocation and promote social cohesion, thereby highlighting its potential role in maintaining colony fitness. Our study shows that the iMScopeTM prototype is a novel AP-MALDI-MSI technique to detect 13C-integrated metabolites in the 13C-labeled sample. This study also demonstrated that this technique can detect 13C-integrated PE, which is abundant mainly in termite abdomen. Mini-abstract.